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Methicillin-resistant <i>Staphylococcus aureus</i> (MRSA) is a serious pathogen in clinical settings and early detection is critical. Here, we investigated the MRSA discrimination potential of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) using 320 clinical <i>S. aureus</i> isolates obtained in 2005&#8722;2014 and 181 isolates obtained in 2018. We conducted polymerase chain reactions (PCR) for staphylococcal cassette chromosome <i>mec</i> (SCC<i>mec</i>) typing and MALDI-TOF MS to find specific markers for methicillin resistance. We identified 21 peaks with significant differences between MRSA and methicillin-susceptible <i>S. aureus</i> (MSSA), as determined by <i>mecA</i> and SCC<i>mec</i> types. Each specific peak was sufficient to discriminate MRSA. We developed two methods for simple discrimination according to these peaks. First, a decision tree for MRSA based on six MRSA-specific peaks, three MSSA-specific peaks, and two SCC<i>mec</i> type IV peaks showed a sensitivity of 96.5%. Second, simple discrimination based on four MRSA-specific peaks and one MSSA peak had a maximum sensitivity of 88.3%. The decision tree applied to 181 <i>S. aureus</i> isolates from 2018 had a sensitivity of 87.6%. In conclusion, we used specific peaks to develop sensitive MRSA identification methods. This rapid and easy MALDI-TOF MS approach can improve patient management.